巢式PCR扩增孕妇血浆中胎儿ZFY基因的研究

The study of fetal DNA in maternal plasma using the nested polymerase chain reaction

目的:建立一种无创伤性产前基因诊断的方法。方法:自行建立巢式PCR方法对46例10～40孕周 的初孕妇血浆中游离胎儿Y染色体ZFY基因进行扩增,扩增片段分别为354bp和307bp,46例均采用母体血浆直 接作为模板进行巢式PCR。结果:39例妊娠男性胎儿孕妇血浆中有32例出现ZFY基因条带,检出率为82.1% (32/39),其中24例2次扩增均为阳性,8例第2次扩增才出现阳性条带,巢式PCR可明显提高检测敏感性(从 61.5%提高到82.1%)。7例妊娠女性胎儿孕妇血浆2次均未出现阳性扩增带,无假阳性结果。本研究的性别总 符合率为84.8%(39/46)。结论:巢式聚合酶链反应检测母体血浆中游离胎儿DNA的敏感性和特异性较高,有希 望成为无创伤性产前基因诊断方法应用于临床。

Objective:To establish a new method for non-invasive prenatal diagnosis. Methods:A single-copy ZFY gene of Y-chromosome of fetal DNA was amplified by nested polymerase chain reaction (PCR) from 46 pregnant maternal plasma (10-40 weeks) . A 354 bp and 307 bp specific fragment were obtained. The maternal plasma samples of 46 pregnant women were used directly for nested PCR. Results:The fragment was identified in 32 of 39 pregnant women plasma bearing male fetuses. The diagnostic rate was 82.1%(32/39), 24 of 32 women have positive band in both amplifications, 8 of 32 women have positive band at the second amplification. The positive rate was increased by nested PCR (from 61.5% to 82.1%). The total accuracy was 84.8%(39/46). Conclusion:The circulating fetal DNA in maternal plasma may have been detected by nested PCR which have high sensitivity and specificity for the clinical application.